This invention relates to novel multibinding compounds (agents) that are muscarinic receptor antagonists, pharmaceutical compositions comprising such compounds, and methods of preparing these compounds. Accordingly, the multibinding compounds and pharmaceutical compositions of this invention are useful in the treatment and prevention of diseases mediated by these receptors such as chronic obstructive pulmonary disease, chronic bronchitis, irritable bowel syndrome, urinary incontinence, and the like.
The following publications are cited in this application as superscript numbers:
1 Bonner, T. I. et al., Science (Washington D.C.) 1987, 237, 527-532.
2 Goyal, R. K., J. Med, 1989, 321, 1022.
3 Hulme, E. C., et al., Annu. Rev. Pharmacol. Toxicol. 1990, 30, 633.
4 Eglen, R. M. and Hegde, S. S., Drug News Perspect. 1997, 10(8), 462-469.
5 Fisher, A., Invest. Drugs, 1997, 6(10), 1395-1411.
6 Martel, A. M., et al., Drugs Future, 1997, 22(2), 135-137.
7 Graul, A. and Castaner, J., Drugs Future, 1996, 21(11), 1105-1108.
8 Graul, A., et al., Drugs Future, 1997, 22(7), 733-737.
All of the above publications are herein incorporated by reference in their entirety to the same extent as if each individual publication was specifically and individually indicated to be incorporated by reference in its entirety.
A receptor is a biological structure with one or more binding domains that reversibly complexes with one or more ligands, where that complexation has biological consequences. Receptors can exist entirely outside the cell (extracellular receptors), within the cell membrane (but presenting sections of the receptor to the extracellular milieu and cytosol), or entirely within the cell (intracellular receptors). They may also function independently of a cell (e.g., clot formation). Receptors within the cell membrane allow a cell to communicate with the space outside of its boundaries (i.e., signaling) as well as to function in the transport of molecules and ions into and out of the cell.
A ligand is a binding partner for a specific receptor or family of receptors. A ligand may be the endogenous ligand for the receptor or alternatively may be a synthetic ligand for the receptor such as a drug, a drug candidate or a pharmacological tool.
The super family of seven transmembrane proteins (7-TMs), also called G-protein coupled receptors (GPCRs), represents one of the most significant classes of membrane bound receptors that communicate changes that occur outside of the cell""s boundaries to its interior, triggering a cellular response when appropriate. The G-proteins, when activated, affect a wide range of downstream effector systems both positively and negatively (e.g., ion channels, protein kinase cascades, transcription, transmigration of adhesion proteins, and the like).
Muscarinic receptors are members of the G-protein coupled receptors that are composed of a family of five receptor sub-types (M1, M2, M3, M4 and M5) and are activated by the neurotransmitter acetylcholine1. These receptors are widely distributed on multiple organs and tissues and are critical to the maintenance of central and peripheral cholinergic neurotransmission. The regional distribution of these receptor subtypes in the brain and other organs has been documented1-4. For example, the smooth muscle is composed largely of M2 and M3 receptors, cardiac muscle is composed largely of M2 receptors, and salivary glands are largely, composed of M3 receptors.
It has been established that the muscarinic receptors are involved in diseases such as chronic obstructive pulmonary disease5-6, asthma, irritable bowel syndrome7, urinary incontinence7-8, rhinitis, spasmodic colitis, chronic cystitis, and alzheimer,s disease, senile dementia, glaucoma, schizophrenia, gastroesophogeal reflux disease, cardiac arrhythmia, and hyper salvation syndromes. Currently, a number of compounds having muscarinic receptor antagonistic activities are being used to treat these diseases. For example, oxybutynin is being used for the treatment of urinary urge incontinence and dicyclomine for the treatment of irritable bowel syndrome. However, these drugs have limited utility as they produce side effects such as dry mouth, blurred vision, and mydriasis. Therefore, there is a need for muscarinic receptor antagonists that will help in the treatment of the above diseases without the adverse side effects.
The multibinding compounds of the present invention fulfill this need.
This invention is directed to novel multibinding compounds (agents) that are muscarinic receptor antagonists and are therefore useful in the treatment and prevention of diseases such as chronic obstructive pulmonary disease, chronic bronchitis, irritable bowel syndrome, urinary incontinence, and the like.
Accordingly, in one of its composition aspects, this invention provides a multibinding compound comprising of from 2 to 10 ligands covalently attached to one or more linkers, wherein each of said ligands comprises, independently of each other, a muscarinic receptor antagonist or an allosteric modulator of a muscarinic receptor, and pharmaceutically acceptable salts thereof provided that at least one of said ligands is a muscarinic receptor antagonist and further provided that when the multibinding compound comprises 2 or 3 ligands, then only one of the ligands is 11-acetyl-5,11-dihydro-6H-pyrido[2,3b][1,4]benzodiazepin-6-one, N-methylquinuclidine, or a compound of formula: 
wherein:
na is 0 or 1;
Rc is hydrogen or alkyl;
Rd is hydrogen; and
Re is xe2x80x94CO2CRf(phenyl)2 wherein Rf is hydrogen or hydroxy.
In a second aspect, this invention provides a multibinding compound of Formula (I):
(L)p(X)qxe2x80x83xe2x80x83(I)
wherein:
each L is, independently of each other, a muscarinic receptor antagonist or an allosteric modulator of a muscarinic receptor;
each X is independently a linker;
p is an integer of from 2 to 10; and
q is an integer of from 1 to 20, and pharmaceutically acceptable salts thereof, provided that at least one of said ligands is a muscarinic receptor antagonist, and further provided that when the multibinding compound comprises of 2 or 3 ligands, then only one of the ligands is 11-acetyl-5,11-dihydro-6H-pyrido[2,3][1,4]benzodiazepin-6-one, N-methylquinuclidine, or a compound of formula: 
wherein:
na is 0 or 1;
Rc is hydrogen or alkyl;
Rd is hydrogen; and
Re is xe2x80x94CO2CRf(phenyl)2 wherein Rf is hydrogen or hydroxy.
Preferably, q is less than p in the multibinding compounds of this invention.
Preferably, each ligand, L, that is a muscarinic receptor antagonist in the multibinding compound of Formula (I) is independently selected from the group consisting of:
(1) A Compound of Formula (a): 
wherein:
A is an aryl or a heteroaryl ring;
Bxe2x80x3 is xe2x80x94CH2xe2x80x94, xe2x80x94Oxe2x80x94 or xe2x80x94NRaxe2x80x94 where Ra is hydrogen, alkyl, or substituted alkyl;
R1 is hydrogen or alkyl;
R2 is selected from a group consisting of formula (i), (ii), (iii), or xe2x80x9cHetxe2x80x9d: 
wherein:
xe2x80x94xe2x80x94 is an optional double bond;
n1 is an integer of from 1 to 4;
n2 is an integer of from 1 to 3;
V is xe2x80x94CHxe2x80x94, xe2x80x94Oxe2x80x94, xe2x80x94S(O)n3xe2x80x94 (where n3 is an integer of from 0 to 2), or xe2x80x94NR4xe2x80x94 (wherein R4 is hydrogen, alkyl, substituted alkyl, aryl, or heteroaryl);
xe2x80x9cHetxe2x80x9d is a heteroaryl ring which optionally attaches the ligand to a linker;
R3 is hydrogen, alkyl, amino, substituted amino, xe2x80x94OR3 (where Ra is hydrogen, alkyl, or acyl), or a covalent bond attaching the ligand to a linker;
R5 is hydrogen, alkyl, amino, substituted amino, xe2x80x94ORb (where Rb is hydrogen or alkyl), aryl, aralkyl, heteroaralkyl, or a covalent bond attaching the ligand to a linker;
R6, R7, and R8 are, independently of each other, hydrogen, halo, hydroxy, alkoxy, haloalkoxy, carboxy, alkoxycarbonyl, alkyl optionally substituted with one, two or three substituents selected from halo, hydroxy, carboxy, alkoxycarbonyl, alkylthio, alkylsulfonyl, amino, substituted amino, or a covalent bond attaching the ligand to a linker;
K is a bond or an alkylene group;
Kxe2x80x3 is a bond, xe2x80x94C(O)xe2x80x94, xe2x80x94S(O)n4xe2x80x94 (where n4 is an integer of from 0 to 2), or an alkylene group optionally substituted with a hydroxyl group; and
B is a heterocycloamino group which optionally attaches the ligand to a linker;
provided that at least one of the R5, R6, R7, R8, xe2x80x9cHetxe2x80x9d, or the heterocycloamino group attaches the ligand to a linker;
(2) A Compound of Formula (b): 
wherein:
C is an aryl or heteroaryl ring which optionally attaches the ligand to a linker;
R9 is hydrogen, hydroxy, cyano, aminocarbonyl which optionally links the ligand to a linker, alkyl optionally substituted with one, two or three substituents selected from halo, hydroxy, carboxy, alkoxycarbonyl, alkylthio, alkylsulfonyl, amino, substituted amino, or a covalent bond attaching the ligand to a linker;
R10 is hydrogen, aryl, heteroaryl, cycloalkyl, alkyl optionally substituted with one, two or three substituents selected from halo, hydroxy, carboxy, alkoxycarbonyl, alkylthio, alkylsulfonyl, amino, substituted amino, or a covalent bond attaching the ligand to a linker;
Q is a single bond, xe2x80x94Oxe2x80x94, xe2x80x94COCH2xe2x80x94, xe2x80x94C(O)NHxe2x80x94, xe2x80x94NHC(O)Oxe2x80x94, xe2x80x94NHC(O)NHxe2x80x94, or xe2x80x94C(O)Oxe2x80x94;
Qxe2x80x3 is selected from the group consisting of:
(i) monoaalkylaminoalkyl, monoalkylaminoalkenyl, monoalkylaminoalkynyl wherein the amino group optionally links the ligand to a linker;
(ii) carboxy which optionally links the ligand to a linker;
(iii) a group of formula (iv): 
where:
E is hydrogen, a covalent bond attaching the ligand to a linker, or xe2x80x94CH2xe2x80x94Wxe2x80x94 R11 wherein W is a single bond or alkylene wherein one of the carbon atoms may optionally be replaced by xe2x80x94Oxe2x80x94, xe2x80x94Sxe2x80x94, or xe2x80x94NRgxe2x80x94 (wherein Rg is hydrogen or alkyl); and
R11 is a group of formula (v), (vi), or xe2x80x9cHetxe2x80x9d: 
wherein:
xe2x80x94 is an optional bond;
T and U are, independently of each other, xe2x80x94Oxe2x80x94 or xe2x80x94CH2xe2x80x94;
n5 is an integer of from 1 to 3; and
xe2x80x9cHetxe2x80x9d is heteroaryl; and
(iv) a group of formula (vii), (viii) or (ix): 
wherein:
n6 is 0 or 1;
Mxe2x88x92 is a counterion;
R12 is a covalent bond attaching the ligand to a linker;
R13 is alkyl, alkenyl, cycloalkyl, or a covalent bond attaching the ligand to a linker;
R14 is hydrogen, alkyl, or a covalent bond attaching the ligand to a linker;
R51 is hydrogen or alkyl; and
J is:
xe2x80x94(CH2)2xe2x80x94, xe2x80x94(CH2)3xe2x80x94, or

provided that at least one of the C, R9, R10, and Qxe2x80x3 attaches the ligand to a linker;
(3) A Compound of Formula (c): 
wherein:
Gxe2x80x2 is pyrrolidine, piperidine, or 
wherein said Gxe2x80x2 groups optionally attach the ligand to a linker;
n7 is an 0 or 1, provided that when the nitrogen atom of the quinclidine ring attaches the ligand to the linker then n7 is 0;
n8 is 1 or 2;
g is an integer of from 0 to 3;
each R15 is, independently of each other, hydrogen, halogen, nitro, cyano, hydroxy, alkoxy, carboxy, alkoxycarbonyl, acyl, thio, alkylthio, alkylsulfonyl, alkylsulfinyl, sulfonamido, alkylsulfonamido, carbamoyl, thiocarbamoyl, mono or dialkylcarbamoyl, amino, mono- or dialkylamino, methylenedioxy, ethylenedioxy, alkyl optionally substituted with one, two or three substituents selected from halo, hydroxy, carboxy, alkoxycarbonyl, alkylthio, alkylsulfonyl, amino, or substituted amino, or a covalent bond attaching the ligand to a linker;
G is aryl, heteroaryl, heterocyclyl, or cycloalkyl which optionally attach the ligand to a linker; and
Gxe2x80x3 is a single bond or an alkylene group provided that at least one of the R15, G, Gxe2x80x2, and Gxe2x80x3 attaches the ligand to a linker;
(4) A Compound of Formula (d): 
wherein:
n9 is 1 or 2;
n10 is 0 or 1 provided that n9+n10 is 1 or 2;
P is an aryl or heteroaryl ring which optionally attaches the ligand to a linker;
Pxe2x80x3 is a single bond or an alkylene group;
S is a heterocycloamino ring which optionally attaches the ligand to a linker provided that at least one of the P and S attaches the ligand to a linker;
(5) A Compound of Formula (e): 
wherein:
n14 is 0, 1, or 2;
n52 is 0 or 1;
R16 is hydrogen, alkyl, or a covalent bond attaching the ligand to a linker;
R17, R18, and R19 are, independently of each other, hydrogen, alkyl, alkoxy, hydroxy, carbamoyl, sulfanoyl, halo, or a covalent bond attaching the ligand to a linker;
R20 and R21 are, independently of each other, hydrogen, alkyl or a covalent bond attaching the ligand to a linker; or R20 and R21 together with the nitrogen atom to which they are attached form a heterocycloamino ring which optionally attaches the ligand to a linker provided that at least one of the R16, R17, R18, R19, R20, and R21 attaches the ligand to a linker; or
(6) A Compound of Formula (f): 
wherein:
R22 is hydrogen or halo;
R23 is alkyl, cycloalkyl, aryl, heteroaryl, heterocyclyl, or heterocyclylalkyl;
R24 is hydroxy, halo, or a covalent bond attaching the ligand to a linker;
R25 and R26 are, independently of each other, hydrogen, alkyl, aralkyl, or a covalent bond attaching the ligand to a linker, or R25 and R26 together with the nitrogen atom to which they are attached form a heterocycloamino group which optionally attaches the ligand to a linker provided that at least one of the R24, R25, and R26 attaches the ligand to a linker; and
each ligand, L, that is an allosteric modulator of a muscarinic receptor in the multibinding compound of Formula (I) is independently selected from a group consisting of:
(7) A Compound of Formula (g): 
wherein:
Dxe2x80x3 is alkylene;
D is xe2x80x94NR31R32, xe2x80x94N+(R33R34R35) or xe2x80x94OR32 where R31, R33, and R34 are, independently of each other, hydrogen, alkyl, or aralkyl; and R32 and R35 represent a covalent bond attaching the ligand to a linker;
R27 is hydrogen, halo, nitro, cyano, hydroxy, alkoxy, carboxy, alkoxycarbonyl, acyl, thio, alkylthio, alkylsulfonyl, alkylsulfinyl, sulfonamido, alkylsulfonamido, carbamoyl, thiocarbamoyl, mono or dialkylcarbamoyl, amino, mono- or dialkylamino, aryl, aryloxy, arylthio, heteroaryl, heteraryloxy, heteroarylthio, heterocyclyl, heterocyclyloxy, aralkyl, heteroaralkyl, or alkyl optionally substituted with one, two or three substituents selected from halo, hydroxy, carboxy, alkoxycarbonyl, alkylthio, alkylsulfonyl, amino, or substituted amino;
R28 is hydrogen, halo, nitro, cyano, hydroxy, alkoxy, carboxy, alkoxycarbonyl, acyl, thio, alkylthio, alkylsulfonyl, alkylsulfinyl, sulfonamido, alkylsulfonamido, carbamoyl, thiocarbamoyl, mono or dialkylcarbamoyl, amino, mono- or dialkylamino, or alkyl optionally substituted with one, two, or three substituents selected from halo, hydroxy, carboxy, alkoxycarbonyl, alkylthio, alkylsulfonyl, amino, or substituted amino;
R29 and R30 are, independently of each other, hydrogen, alkyl, haloalkyl, halo, nitro, cyano, hydroxy, alkoxy, alkoxycarbonyl, acyl, thio, alkylthio, amino, mono- or dialkylamino; or
one of R27, R28, R29, or R30 together with the adjacent group forms a methylenedioxy or ethylenedioxy group;
(8) A Compound of Formula (h): 
wherein:
n11 is an integer of from 1 to 7;
n12 is 0 to 7;
F is xe2x80x94NR40xe2x80x94, xe2x80x94Oxe2x80x94, xe2x80x94Sxe2x80x94, or xe2x80x94CHR41xe2x80x94 (wherein R40 and R41 are, independently of each other, hydrogen, alkyl, or substituted alkyl);
Fxe2x80x2 is a covalent bond, xe2x80x94OR43, xe2x80x94NR42R43 or xe2x80x94N+R43R44R45 wherein R42 is hydrogen or alkyl, R44 and R45 are alkyl, and R43 is a covalent bond attaching the ligand to a linker;
R36 is hydrogen, alkyl, halo, nitro, cyano, hydroxy, alkoxy, carboxy, alkoxycarbonyl, acyl, thio, alkylthio, alkylsulfonyl, alkylsulfinyl, sulfonamido, alkylsulfonamido, carbamoyl, thiocarbamoyl, mono or dialkylcarbamoyl, amino, mono- or dialkylamino, aryl, aryloxy, arylthio, heteroaryl, heteraryloxy, heteroarylthio, heterocyclyl, heterocyclyloxy, aralkyl, heteroaralkyl, or alkyl optionally substituted with one, two or three substituents selected from halo, hydroxy, carboxy, alkoxycarbonyl, alkylthio, alkylsulfonyl, amino, or substituted amino;
R37 is hydrogen, alkyl, halo, nitro, cyano, hydroxy, alkoxy, alkoxycarbonyl, acyl, thio, alkylthio, amino, mono- or dialkylamino, aryl, aryloxy, arylthio, heteroaryl, heteraryloxy, heteroarylthio, heterocyclyl, heterocyclyloxy, aralkyl, heteroaralkyl, or alkyl optionally substituted with one, two or three substituents selected from halo, hydroxy, carboxy, alkoxycarbonyl, alkylthio, alkylsulfonyl, amino, or substituted amino; and
R38 is hydrogen, alkyl, halo, hydroxy, alkoxy, or a covalent bond attaching the ligand to a linker provided that at least one of R38 and R43 attaches the ligand to a linker;
R39 is hydrogen, alkyl, halo, hydroxy, alkoxy, or substituted alkyl; or
(9) A Compound of Formula (i): 
wherein:
R46 is alkyl, substituted alkyl, cycloalkyl, substituted cycloalkyl, or heterocycle;
R47 is alkyl, substituted alkyl, aryl, acyl, heterocycle, or xe2x80x94COOR49 where R49 is alkyl; or
R46 and R47 together with the nitrogen atom to which they are attached form heterocycle;
R48 is a covalent bond that attaches the ligand to a linker;
R49 is alkyl; and pharmaceutically acceptable salts, individual isomers, mixture of isomers, and prodrugs thereof provided that at least one of the ligands is a muscarinic receptor antagonist.
Preferably, each ligand, L, that is a muscarinic receptor antagonist in the multibinding compound of Formula (I) is independently selected from a group consisting of Darifenacin, Tolterodine, Oxybutynin, YM-46303, YM-58790, 5-(2-isopropylimidazol-1-yl)-3,3-diphenyl-2(3H)furanone, 5-(imidazol-1-ylmethyl)-3,3-diphenyl-2(3H)furanone which is linked to a linker at the 2-position of imidazole ring, 5-(N-ethylaminomethyl)-3,3-diphenyl-2(3H)furanone which is linked to a linker via the secondary amino group, 3,3-diphenyl-2(3H)furanone which is linked to a linker at the 5-position of the furanone ring (disclosed in J. Med. Chem., 35, 4415-4424, 1992), 3-[4-(2-chlorobenzyl)piperazin-1-yl)-1-cyclobutyl-1-phenyl-2-propanone which is linked to a linker via the phenyl ring of the benzyl moiety, 3-(piperazin-1-yl)-1-cyclobutyl-1-phenyl-2-propanone which is linked to a linker via the piperazine ring, 3-[4-(benzylpiperazin-1-yl)-1-cyclobutyl-1-phenyl-2-propanone which is linked to a linker via the phenyl ring of the benzyl moiety, 3-[4-benzylpiperazin-1-yl)-1,1-diphenyl-2-propanone which is linked to a linker via the phenyl ring of the benzyl moiety, 3-(piperazin-1-yl)-1,1-diphenyl-2-propanone which is linked to a linker via the piperazine ring (disclosed in J. Med. Chem., 36, 610-616, 1993), the derivatives thereof.
Preferably, each linker, X, in the multibinding compound of Formula (I) independently has the formula:
xe2x80x94Xaxe2x80x94Zxe2x80x94(Yaxe2x80x94Z)mxe2x80x94Ybxe2x80x94Zxe2x80x94Xaxe2x80x94
wherein
m is an integer of from 0 to 20;
Xa at each separate occurrence is selected from the group consisting of xe2x80x94Oxe2x80x94, xe2x80x94Sxe2x80x94, xe2x80x94NRxe2x80x94, xe2x80x94C(O)xe2x80x94, xe2x80x94C(O)Oxe2x80x94, xe2x80x94C(O)NRxe2x80x94, xe2x80x94C(S), xe2x80x94C(S)Oxe2x80x94, xe2x80x94C(S)NRxe2x80x94 or a covalent bond where R is as defined below;
Z at each separate occurrence is selected from the group consisting of alkylene, substituted alkylene, cycloalkylene, substituted cylcoalkylene, alkenylene, substituted alkenylene, alkynylene, substituted alkynylene, cycloalkenylene, substituted cycloalkenylene, arylene, heteroarylene, heterocyclene, or a covalent bond;
Ya and Yb at each separate occurrence are selected from the group consisting of xe2x80x94Oxe2x80x94, xe2x80x94C(O)xe2x80x94, xe2x80x94OC(O)xe2x80x94, xe2x80x94C(O)Oxe2x80x94, xe2x80x94NRxe2x80x94, xe2x80x94S(O)nxe2x80x94, xe2x80x94C(O)NRxe2x80x2xe2x80x94, xe2x80x94NRxe2x80x2C(O)xe2x80x94, xe2x80x94NRxe2x80x2C(O)NRxe2x80x2xe2x80x94, xe2x80x94NRxe2x80x2C(S)NRxe2x80x2xe2x80x94, xe2x80x94C(xe2x95x90NRxe2x80x2)xe2x80x94NRxe2x80x2xe2x80x94, xe2x80x94NRxe2x80x2xe2x80x94C(xe2x95x90NRxe2x80x2)xe2x80x94, xe2x80x94OC(O)xe2x80x94NRxe2x80x2xe2x80x94, xe2x80x94NRxe2x80x2xe2x80x94C(O)xe2x80x94Oxe2x80x94, xe2x80x94Nxe2x95x90C(Xa)xe2x80x94NRxe2x80x2xe2x80x94, xe2x80x94NRxe2x80x2xe2x80x94C(Xa)xe2x95x90Nxe2x80x94, xe2x80x94P(O)(ORxe2x80x2)xe2x80x94Oxe2x80x94, xe2x80x94Oxe2x80x94P(O)(ORxe2x80x2)xe2x80x94, xe2x80x94S(O)nCRxe2x80x2Rxe2x80x3xe2x80x94, xe2x80x94S(O)nxe2x80x94NRxe2x80x2xe2x80x94, xe2x80x94NRxe2x80x2xe2x80x94S(O)nxe2x80x94, xe2x80x94Sxe2x80x94Sxe2x80x94, and a covalent bond; where n is 0, 1 or 2; and R, Rxe2x80x2 and Rxe2x80x3 at each separate occurrence are selected from the group consisting of hydrogen, alkyl, substituted alkyl, cycloalkyl, substituted cycloalkyl, alkenyl, substituted alkenyl, cycloalkenyl, substituted cycloalkenyl, alkynyl, substituted alkynyl, aryl, heteroaryl and heterocyclic.
In a third aspect, this invention provides a pharmaceutical composition comprising a pharmaceutically acceptable carrier and an effective amount of a multibinding compound comprising from 2 to 10 ligands covalently attached to one or more linkers, wherein each of said ligands wherein each of said ligands comprises, independently of each other, a muscarinic receptor antagonist or an allosteric modulator of a muscarinic receptor provided that at least one of said ligands is a muscarinic receptor antagonist, and pharmaceutically acceptable salts thereof.
In a fourth aspect, this invention provides a method of treating diseases mediated by a muscarinic receptor in a mammal, said method comprising administering to said mammal a therapeutically effective amount of a pharmaceutical composition comprising a pharmaceutically acceptable carrier and a multibinding compound comprising from 2 to 10 ligands covalently attached to one or more linkers, wherein each of said ligands, comprises, independently of each other, a muscarinic receptor antagonist or an allosteric modulator of a muscarinic receptor provided that at least one of said ligands is a muscarinic receptor antagonist, and pharmaceutically acceptable salts thereof.
In a fifth aspect, this invention is directed to general synthetic methods for generating large libraries of diverse multimeric compounds which multimeric compounds are candidates for possessing multibinding properties for a muscarinic receptor. The diverse multimeric compound libraries provided by this invention are synthesized by combining a linker or linkers with a ligand or ligands to provide for a library of multimeric compounds wherein the linker and ligand each have complementary functional groups permitting covalent linkage. The library of linkers is preferably selected to have diverse properties such as valency, linker length, linker geometry and rigidity, hydrophilicity or hydrophobicity, amphiphilicity, acidity, basicity and polarization. The library of ligands is preferably selected to have diverse attachment points on the same ligand, different functional groups at the same site of otherwise the same ligand, and the like.
This invention is also directed to libraries of diverse multimeric compounds which multimeric compounds are candidates for possessing multibinding properties for a muscarinic receptor. These libraries are prepared via the methods described above and permit the rapid and efficient evaluation of what molecular constraints impart multibinding properties to a ligand or a class of ligands targeting a muscarinic receptor.
Accordingly, in one of its method aspects, this invention is directed to a method for identifying multimeric ligand compounds possessing multibinding properties for a muscarinic receptor which method comprises:
(a) identifying a ligand or a mixture of ligands wherein each ligand contains at least one reactive functionality;
(b) identifying a library of linkers wherein each linker in said library comprises at least two functional groups having complementary reactivity to at least one of the reactive functional groups of the ligand;
(c) preparing a multimeric ligand compound library by combining at least two stoichiometric equivalents of the ligand or mixture of ligands identified in (a) with the library of linkers identified in (b) under conditions wherein the complementary functional groups react to form a covalent linkage between said linker and at least two of said ligands; and
(d) assaying the multimeric ligand compounds produced in (c) above to identify multimeric ligand compounds possessing multibinding properties for a muscarinic receptor.
In another of its method aspects, this invention is directed to a method for identifying multimeric ligand compounds possessing multibinding properties for a muscarinic receptor which method comprises:
(a) identifying a library of ligands wherein each ligand contains at least one reactive functionality;
(b) identifying a linker or mixture of linkers wherein each linker comprises at least two functional groups having complementary reactivity to at least one of the reactive functional groups of the ligand;
(c) preparing a multimeric ligand compound library by combining at least two stoichiometric equivalents of the library of ligands identified in (a) with the linker or mixture of linkers identified in (b) under conditions wherein the complementary functional groups react to form a covalent linkage between said linker and at least two of said ligands; and
(d) assaying the multimeric ligand compounds produced in (c) above to identify multimeric ligand compounds possessing multibinding properties for a muscarinic receptor.
The preparation of the multimeric ligand compound library is achieved by either the sequential or concurrent combination of the two or more stoichiometric equivalents of the ligands identified in (a) with the linkers identified in (b). Sequential addition is preferred when a mixture of different ligands is employed to ensure heterodimeric or multimeric compounds are prepared. Concurrent addition of the ligands occurs when at least a portion of the multimer compounds prepared are homomultimeric compounds.
The assay protocols recited in (d) can be conducted on the multimeric ligand compound library produced in (c) above, or preferably, each member of the library is isolated by preparative liquid chromatography mass spectrometry (LCMS).
In one of its composition aspects, this invention is directed to a library of multimeric ligand compounds which may possess multivalent properties for a muscarinic receptor which library is prepared by the method comprising:
(a) identifying a ligand or a mixture of ligands wherein each ligand contains at least one reactive functionality;
(b) identifying a library of linkers wherein each linker in said library comprises at least two functional groups having complementary reactivity to at least one of the reactive functional groups of the ligand; and
(c) preparing a multimeric ligand compound library by combining at least two stoichiometric equivalents of the ligand or mixture of ligands identified in (a) with the library of linkers identified in (b) under conditions wherein the complementary functional groups react to form a covalent linkage between said linker and at least two of said ligands.
In another of its composition aspects, this invention is directed to a library of multimeric ligand compounds which may possess multivalent properties for a muscarinic receptor which library is prepared by the method comprising:
(a) identifying a library of ligands wherein each ligand contains at least one reactive functionality;
(b) identifying a linker or mixture of linkers wherein each linker comprises at least two functional groups having complementary reactivity to at least one of the reactive functional groups of the ligand; and
(c) preparing a multimeric ligand compound library by combining at least two stoichiometric equivalents of the library of ligands identified in (a) with the linker or mixture of linkers identified in (b) under conditions wherein the complementary functional groups react to form a covalent linkage between said linker and at least two of said ligands.
In a preferred embodiment, the library of linkers employed in either the methods or the library aspects of this invention is selected from the group comprising flexible linkers, rigid linkers, hydrophobic linkers, hydrophilic linkers, linkers of different geometry, acidic linkers, basic linkers, linkers of different polarization and amphiphilic linkers. For example, in one embodiment, each of the linkers in the linker library may comprise linkers of different chain length and/or having different complementary reactive groups. Such linker lengths can preferably range from about 2 to 100 xc3x85, more preferably 2-25xc2x0 A.
In another preferred embodiment, the ligand or mixture of ligands is selected to have reactive functionality at different sites on said ligands in order to provide for a range of orientations of said ligand on said multimeric ligand compounds. Such reactive functionality includes, by way of example, carboxylic acids, carboxylic acid halides, carboxyl esters, amines, halides, isocyanates, vinyl unsaturation, ketones, aldehydes, thiols, alcohols, anhydrides, and precursors thereof. It is understood, of course, that the reactive functionality on the ligand is selected to be complementary to at least one of the reactive groups on the linker so that a covalent linkage can be formed between the linker and the ligand.
In other embodiments, the multimeric ligand compound is homomeric (i.e., each of the ligands is the same, although it may be attached at different points) or heterodimeric (i.e., at least one of the ligands is different from the other ligands).
In addition to the combinatorial methods described herein, this invention provides for an interative process for rationally evaluating what molecular constraints impart multibinding properties to a class of multimeric compounds or ligands targeting a muscarinic receptor. Specifically, this method aspect is directed to a method for identifying multimeric ligand compounds possessing multibinding properties for a muscarinic receptor which method comprises:
(a) preparing a first collection or iteration of multimeric compounds which is prepared by contacting at least two stoichiometric equivalents of the ligand or mixture of ligands which target a receptor with a linker or mixture of linkers wherein said ligand or mixture of ligands comprises at least one reactive functionality and said linker or mixture of linkers comprises at least two functional groups having complementary reactivity to at least one of the reactive functional groups of the ligand wherein said contacting is conducted under conditions wherein the complementary functional groups react to form a covalent linkage between said linker and at least two of said ligands;
(b) assaying said first collection or iteration of multimeric compounds to assess which if any of said multimeric compounds possess multibinding properties for a muscarinic receptor;
(c) repeating the process of (a) and (b) above until at least one multimeric compound is found to possess multibinding properties for a muscarinic receptor;
(d) evaluating what molecular constraints imparted multibinding properties to the multimeric compound or compounds found in the first iteration recited in (a)-(c) above;
(e) creating a second collection or iteration of multimeric compounds which elaborates upon the particular molecular constraints imparting multibinding properties to the multimeric compound or compounds found in said first iteration;
(f) evaluating what molecular constraints imparted enhanced multibinding properties to the multimeric compound or compounds found in the second collection or iteration recited in (e) above;
(g) optionally repeating steps (e) and (f) to further elaborate upon said molecular constraints.
Preferably, steps (e) and (f) are repeated at least two times, more preferably at from 2-50 times, even more preferably from 3 to 50 times, and still more preferably at least 5-50 times.